Transfection of the EGF receptor family members ErbB1,2,3, and 4 into MTLn3 rat mammary adenocarcinoma cells has revealed that ErbB1 and ErbB3 significantly enhance intravasation and metastasis. Based on previous in vivo imaging studies with MTLn3 xenograft tumors, we hypothesize that ErbB1 and ErbB3 enhance intravasation through increased cell polarization and chemotaxis towards blood vessels followed by movement into blood vessels. Endogenous sources of ligands for ErbB1 and ErbB3 are likely to be produced by macrophages at the surface of the invading tumor and around blood vessels. To test this hypothesis we have 3 specific aims. In the first specific aim, in vivo imaging will determine whether tumor cell orientation and movement into blood vessels is increased in ErbB1 and ErbB3 tumors. The roles of PI-3 kinase and ErbBs in intravasation will be tested using specific inhibitors. Finally, the density of macrophages and locations of ErbB ligands in the ErbB1 and ErbB3 overexpressing tumors will be compared to the ErbB2 and empty vector tumors. In Specific Aim 2, we will examine whether transgenic lines carrying mutations in the ErbB2 or PyMT oncogenes and which are defective in metastasis also show defects in cell behavior around blood vessels or macrophage distribution in the primary tumor. In addition, we will test whether ErbB1 and ErbB3 can enhance intravasation in the MMTV-ErbB2 and MMTV-pyMT tumorigenesis models. In Specific Aim 3 we will examine intravasated cells for evidence of activation of the ErbB1 and ErbB3 pathways using both activation specific antibodies and microarray methods.